Structure and biological activities of a new mastoparan isolated from the venom of the hornet Vespa basalis

By gel filtration on a Fractogel TSK HW 50 column followed by cation-exchange chromatography on CM-Trisacryl M, a tetradecapeptide amide, designated ‘mastoparan B’, was purified from the venom of the hornet Vespa basalis. Its amino acid sequence was determined as: Leu-Lys-Leu-Lys-Ser-Ile-Val-Ser-Trp-Ala-Lys-Lys-Val-Leu-NH2 and its molecular mass was measured to be 1611 Da by fast-atom-bombardment mass spectrometry. In addition to having a common structure of vespid mastoparans, the peptide shows a less hydrophobic sequence at positions 1, 2, 5, 8 and 9. The peptide caused liberation of histamine from rat peritoneal mast cells and induced oedema in the rat paw. However, the latter effect was inhibited by ‘anti-serotonin’ (anti-5-hydroxytryptamine) (cyproheptadine), but not by antihistamine (chlorpheniramine). The peptide also possesses a potent haemolytic activity which acts in synergy with the lethal protein of the venom, suggesting the possible involvement of mastoparan B in the lethal effect of Vespa basalis venom.

COMPLEMENT AS A MEDIATOR OF INFLAMMATION

Interaction in free solution of highly purified preparations of human C'1 esterase, C'4, C'2, and C'3, in the presence of Mg2+, resulted in rapid generation of an activity indistinguishable by biological criteria from anaphylatoxin. The formation of anaphylatoxin was associated with immunoelectrophoretic conversion of C'3 to anodically faster migrating proteins and was unaffected by the presence or absence of added C'5. The biological properties of human anaphylatoxin prepared in this manner include: contraction and desensitization of isolated guinea pig ileum, failure to contract isolated rat uterus, enhancement of vascular permeability in guinea pig skin, degranulation of mast cells in guinea pig mesentery preparations, and liberation of histamine from suspensions of rat peritoneal mast cells. The smooth muscle-contracting and permeability enhancing properties were fully blocked by an antihistaminic drug, triprolidine. No cross-desensitizing activity on guinea pig ileum was demonstrable between rat and human or guinea pig and human anaphylatoxins but a closer biological relationship between rat and guinea pig anaphylatoxins was observed. It is concluded that anaphylatoxin is a product of the complement system. Its possible relationship to apparently similar activities currently being obtained in other laboratories has been discussed.

A LABILE SERUM FACTOR IN EXPERIMENTAL ENDOTOXIN SHOCK: CROSS-TRANSFUSION STUDIES IN DOGS

Peripheral vascular failure caused by endotoxin in the dog has an initial stage of vasoconstriction. Preliminary studies in vitro demonstrated that the constriction was due to the interaction of endotoxin with a heat-labile serum or plasma factor and platelets, resulting in the liberation of histamine. Further studies on the intact dog support and extend this concept. A standardized dose of Escherichia coli endotoxin produced fatal shock in control adult mongrel dogs within 28 hours. The characteristic pattern of changes included progressive hypotension, oliguria and anuria, hemoconcentration, and acidosis. Normal dogs were protected against endotoxin by transfusions of blood in which the essential serum factor was depleted in one of two ways. First, plasma separated from the blood of normal animals was heated at 56°C for 30 minutes, and the infused reconstituted whole blood protected normal dogs. Protection was not afforded by unheated reconstituted blood. Second, blood from immune dogs obtained within 24 hours after a second lethal dose of endotoxin protected recipient dogs. However, protection was not demonstrated with blood collected 72 hours after a second injection of endotoxin. The nature of the serum factor essential for endotoxin activity is not known. It is postulated that an enzyme or enzyme system is involved, and the possible role of complement is discussed.

Nature of the Secretory Activity of the Mast Cell

In the tissues of the peritoneal cavity the mast cell appears to be the only cell capable of releasing histamine. Protamine sulfate and toluidine blue in certain concentrations in Tyrode's solution when injected intraperitoneally elicit the liberation of measurable amounts of histamine into the peritoneal fluid. Repeated injections of these substances cause repeated liberation of histamine. No visible changes in the mesenteric mast cells follow this treatment. It is concluded that secretion by the mast cell does not require cell disruption and death as much of the literature indicates but is merocrine in nature. The mast cell appears to be an endocrine cell which can continuously elaborate and release histamine in in response to appropriate stimulation.

Endogenous Histamine Excretion in the Rat as Influenced by X-Ray Irradiation and Compound 48/80

Studies have been made on a spasmogen from rat urine. This material has been identified as histamine. It has been shown that both acute whole body x-ray irradiation and compound 48/80 significantly increase the quantity of endogenous urinary histamine. Endogenous histamine excretion in female rats is approximately 9–10 times greater than in male rats. Neither x-ray irradiation nor compound 48/80 elevate the amount of urinary histamine of the males to that of the female. Histamine liberation is of little or no importance insofar as lethality from acute whole body irradiation is concerned. The amount of histamine liberated by such irradiation is independent of radiation dosage within the range 600–1200 r. After radiation injury, significant levels of urinary histamine were detected only during the first 24 hours. Histamine depletion by chronic administration of compound 48/80 did not prevent further liberation of histamine by acute whole body irradiation. Irradiated animals are much more susceptible to the toxic effects of compound 48/80 than normal animals. Gonadectomy followed by α-estradiol injection did not increase the output of urinary histamine in male rats. Similar treatment of female rats with testosterone did not reduce their urinary histamine output, but a reduction was observed 140 days after surgery. Administration of cortisone acetate to male rats did not increase their excretion of endogenous histamine. Inactivation of diamine oxidase with aminoguanidine had little or no effect on urinary histamine output in male rats, but caused a threefold increase in females. Further elevation in urinary histamine was produced by irradiation or compound 48/80.